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In Vitro Characterization of Unmodified and Pyroglutamylated Alzheimer's Amyloid beta peptide

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Date Issued:
2014
Abstract/Description:
Plaques of amyloid ? peptide (A?) are a hallmark trait of Alzheimer's disease (AD). However, the precise role of A? aggregates is not well understood. Recent studies have identified that naturally occurring N-terminal truncation and pyroglutamylation of A? significantly increases its neurotoxicity by an unknown mechanism. Content of pyroglutamylated A? (pE-A?) in AD brains has been shown to reach up to 50% of total A?. Modified pE-A? co-aggregates with A? by a seeding mechanism and forms structurally distinct and highly toxic oligomers. We studied structural transitions of the full-length A?1-42, its pyroglutamylated form A?pE3-42, their 9:1 (A?1-42/A?pE3-42) and 1:1 molar combinations. Transmission electron microscopy was used to directly visualize the fibrils of the samples in a buffer mimicking physiological environment. Atomic force microscopy measurements were done to determine rate of second nucleation events in fibrils. Thioflavin-T fluorescence indicated that low ionic strength suppressed the aggregation of A?pE3-42 but promoted that of A?1-42, suggesting different paths of fibrillogenesis of unmodified A? and pE-A?. Interestingly, A?pE3-42 at only 10% significantly facilitated the fibrillization of A?1-42 at near physiological ionic strength but had little effect at low salt. Circular dichroism and Fourier transform infrared (FTIR) spectroscopy were used to characterize the structural transitions during fibrillogenesis. In aqueous buffer, both unmodified A? and pE-A? peptides adopted parallel intermolecular ?-structure. Interestingly, A?pE3-42 contained lower ?-sheet content than 13C-A?1-42, while retaining significantly larger fractions of ?-helical and turn structures. Structural details of A? and pE-A? combinations were unveiled by isotope-edited FTIR spectroscopy, using 13C-labeled A?1-42 and unlabeled A?pE3-42. When exposed to environmental humidity, A?pE3-42 not only maintained an increased fraction of ?-helix but also was able to reverse 13C-A?1-42 ?-sheet structure. These data provide a novel structural mechanism for pE-A? hypertoxicity; pE-A? undergoes fasternucleation due to its increased hydrophobicity, thus promoting formation of smaller, hypertoxic oligomers of partial ?-helical structure.
Title: In Vitro Characterization of Unmodified and Pyroglutamylated Alzheimer's Amyloid beta peptide.
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Name(s): Matos, Jason, Author
Tatulian, Suren, Committee Chair
Teter, Kenneth, Committee Member
Davidson, Victor, Committee Member
University of Central Florida, Degree Grantor
Type of Resource: text
Date Issued: 2014
Publisher: University of Central Florida
Language(s): English
Abstract/Description: Plaques of amyloid ? peptide (A?) are a hallmark trait of Alzheimer's disease (AD). However, the precise role of A? aggregates is not well understood. Recent studies have identified that naturally occurring N-terminal truncation and pyroglutamylation of A? significantly increases its neurotoxicity by an unknown mechanism. Content of pyroglutamylated A? (pE-A?) in AD brains has been shown to reach up to 50% of total A?. Modified pE-A? co-aggregates with A? by a seeding mechanism and forms structurally distinct and highly toxic oligomers. We studied structural transitions of the full-length A?1-42, its pyroglutamylated form A?pE3-42, their 9:1 (A?1-42/A?pE3-42) and 1:1 molar combinations. Transmission electron microscopy was used to directly visualize the fibrils of the samples in a buffer mimicking physiological environment. Atomic force microscopy measurements were done to determine rate of second nucleation events in fibrils. Thioflavin-T fluorescence indicated that low ionic strength suppressed the aggregation of A?pE3-42 but promoted that of A?1-42, suggesting different paths of fibrillogenesis of unmodified A? and pE-A?. Interestingly, A?pE3-42 at only 10% significantly facilitated the fibrillization of A?1-42 at near physiological ionic strength but had little effect at low salt. Circular dichroism and Fourier transform infrared (FTIR) spectroscopy were used to characterize the structural transitions during fibrillogenesis. In aqueous buffer, both unmodified A? and pE-A? peptides adopted parallel intermolecular ?-structure. Interestingly, A?pE3-42 contained lower ?-sheet content than 13C-A?1-42, while retaining significantly larger fractions of ?-helical and turn structures. Structural details of A? and pE-A? combinations were unveiled by isotope-edited FTIR spectroscopy, using 13C-labeled A?1-42 and unlabeled A?pE3-42. When exposed to environmental humidity, A?pE3-42 not only maintained an increased fraction of ?-helix but also was able to reverse 13C-A?1-42 ?-sheet structure. These data provide a novel structural mechanism for pE-A? hypertoxicity; pE-A? undergoes fasternucleation due to its increased hydrophobicity, thus promoting formation of smaller, hypertoxic oligomers of partial ?-helical structure.
Identifier: CFE0005378 (IID), ucf:50465 (fedora)
Note(s): 2014-08-01
M.S.
Medicine, Molecular Biology and Micro
Masters
This record was generated from author submitted information.
Subject(s): Amyloid beta -- Alzheimer's Disease -- Amyloid fibrillization -- Isotope edited FTIR -- Thioflavin T -- Pyroglutamylated amyloid beta
Persistent Link to This Record: http://purl.flvc.org/ucf/fd/CFE0005378
Restrictions on Access: public 2014-08-15
Host Institution: UCF

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